321 results on '"Carbohydrate synthesis"'
Search Results
2. Synthesis of the H-type 1 and Lewis B antigens as 6-aminohexyl glycosides.
- Author
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Pickles, Matisse and Auzanneau, France-Isabelle
- Subjects
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GLYCOSIDES , *FUCOSYLATION , *ANTIGENS , *CELL lines , *BROMINE , *GLYCOLS - Abstract
The LebLea heptasaccharide is a tumor-associated carbohydrate antigen that was isolated from the human colonic adenocarcinoma cell line Colo205 and is a good target for the development of anti-cancer immunotherapeutics. However, it displays on its reducing end the Leb tetrasaccharide: α-l-Fucp-(1→2)-β-d-Galp-(1→3)-[α-l-Fucp-(1→4)]-d-GlcNAcp and the H-type 1 (H-1 antigen) trisaccharide: α-l-Fucp-(1→2)-β-d-Galp-(1→3)-d-GlcNAcp that are also found on noncancerous tissues. To discover analogues or fragments of LebLea that could be used as immunotherapeutics while not triggering immune responses against Leb and the H-1 antigen, we have synthesized the Leb tetrasaccharide hexyl glycoside and the Leb and H-1 antigens aminohexyl glycosides to be used in ELISA experiments. We describe an improved preparation of the 6-O-benzyl-2,3,4-tri-O-acetyl-α-d-galactopyranosyl bromide in neutral conditions and demonstrate the importance of appropriately "matching" the reactivity of acceptors with that of glycosyl donors. Mono- and di-fucosylation of a disaccharide diol acceptor with per-benzylated thioethyl fucoside activated in situ with bromine and under halide ion catalysis is described and our results are compared to literature reports. We observed that our fucosylation reactions required higher equivalents of fucosyl donor and extended reaction times than previously reported. We propose that the protecting groups on the galactosyl unit led to a reduced reactivity of the acceptor. The protected intermediates were converted to 6-azido hexyl glycosides and submitted to dissolving metal conditions to give 6-aminohexyl glycosides. We also prepared the n-hexyl tetrasaccharide glycoside Leb that will be used as a soluble antigen in competitive ELISA experiments. [ABSTRACT FROM AUTHOR]
- Published
- 2024
- Full Text
- View/download PDF
3. Bifunctional Azido(thio)ureas from an O-Protected 2-Amino-2-deoxy-d-glucopyranose: Synthesis and Structural Analyses
- Author
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Concepción Sosa-Gil, Esther Matamoros, Pedro Cintas, and Juan C. Palacios
- Subjects
azido(thio)ureas ,amino sugars ,atropisomers ,conformational analysis ,carbohydrate synthesis ,computational chemistry ,Organic chemistry ,QD241-441 - Abstract
This publication reports a facile and convenient preparation of tri-O-acetyl-glucopyranoses, derived from the corresponding 2-deoxyaminosugar, where the vicinal anomeric and C2 positions are decorated by azido and (thio)ureido groups, respectively. This double functionalization leads to an inherently chiral core incorporating the versatile azido and (thio)ureido linkages prone to further manipulation. The latter also provides a structural element for hydrogen-bonded donor-acceptor (HB-DA) sites, which are of immense value in organocatalytic pursuits. A computation-aided conformational analysis unveils the landscape of available conformers and their relative stability. N-aryl (thio)ureas bearing substituents at ortho positions exist as mixtures of M- and P-atropisomeric conformers.
- Published
- 2024
- Full Text
- View/download PDF
4. Strategies for chemoenzymatic synthesis of carbohydrates
- Author
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Li, Wanqing, McArthur, John B, and Chen, Xi
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Biochemistry and Cell Biology ,Biological Sciences ,Medicinal and Biomolecular Chemistry ,Organic Chemistry ,Chemical Sciences ,1.3 Chemical and physical sciences ,Generic health relevance ,Carbohydrate Sequence ,Carbohydrates ,Glycosylation ,Glycosyltransferases ,Synthetic Biology ,Carbohydrate synthesis ,Chemoenzymatic synthesis ,Glycolipid ,Glycosyltransferase ,Regioselective ,Enzyme engineering ,Biochemistry and cell biology ,Organic chemistry - Abstract
Carbohydrates are structurally complex but functionally important biomolecules. Therefore, they have been challenging but attractive synthetic targets. While substantial progress has been made on advancing chemical glycosylation methods, incorporating enzymes into carbohydrate synthetic schemes has become increasingly practical as more carbohydrate biosynthetic and metabolic enzymes as well as their mutants with synthetic application are identified and expressed for preparative and large-scale synthesis. Chemoenzymatic strategies that integrate the flexibility of chemical derivatization with enzyme-catalyzed reactions have been extremely powerful. Briefly summarized here are our experiences on developing one-pot multienzyme (OPME) systems and representative chemoenzymatic strategies from others using glycosyltransferase-catalyzed reactions for synthesizing diverse structures of oligosaccharides, polysaccharides, and glycoconjugates. These strategies allow the synthesis of complex carbohydrates including those containing naturally occurring carbohydrate postglycosylational modifications (PGMs) and non-natural functional groups. By combining these srategies with facile purification schemes, synthetic access to the diverse space of carbohydrate structures can be automated and will not be limited to specialists.
- Published
- 2019
5. Mechanism of [CO 2 ] Enrichment Alleviated Drought Stress in the Roots of Cucumber Seedlings Revealed via Proteomic and Biochemical Analysis.
- Author
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Li, Yiman, Zhang, Wendong, Zhang, Dalong, Zheng, Yinjian, Xu, Yaliang, Liu, Binbin, and Li, Qingming
- Subjects
- *
CUCUMBERS , *DROUGHTS , *CARBON dioxide , *AMINO acid metabolism , *PROTEOMICS , *SECONDARY metabolism , *SEEDLINGS - Abstract
Cucumber is one of the most widely cultivated greenhouse vegetables, and its quality and yield are threatened by drought stress. Studies have shown that carbon dioxide concentration ([CO2]) enrichment can alleviate drought stress in cucumber seedlings; however the mechanism of this [CO2] enrichment effect on root drought stress is not clear. In this study, the effects of different drought stresses (simulated with 0, 5% and 10% PEG 6000, i.e., no, moderate, and severe drought stress) and [CO2] (400 μmol·mol−1 and 800 ± 40 μmol·mol−1) on the cucumber seedling root proteome were analyzed using the tandem mass tag (TMT) quantitative proteomics method. The results showed that after [CO2] enrichment, 346 differentially accumulating proteins (DAPs) were found only under moderate drought stress, 27 DAPs only under severe drought stress, and 34 DAPs under both moderate and severe drought stress. [CO2] enrichment promoted energy metabolism, amino acid metabolism, and secondary metabolism, induced the expression of proteins related to root cell wall and cytoskeleton metabolism, effectively maintained the balance of protein processing and degradation, and enhanced the cell wall regulation ability. However, the extent to which [CO2] enrichment alleviated drought stress in cucumber seedling roots was limited under severe drought stress, which may be due to excessive damage to the seedlings. [ABSTRACT FROM AUTHOR]
- Published
- 2022
- Full Text
- View/download PDF
6. β-Glucan phosphorylases in carbohydrate synthesis.
- Author
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Ubiparip, Zorica, De Doncker, Marc, Beerens, Koen, Franceus, Jorick, and Desmet, Tom
- Subjects
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CARBOHYDRATES , *PHOSPHORYLASES , *GLUCANS , *BETA-glucans , *DEGREE of polymerization , *SUGAR phosphates , *BIOACTIVE compounds , *POLYMERS - Abstract
β-Glucan phosphorylases are carbohydrate-active enzymes that catalyze the reversible degradation of β-linked glucose polymers, with outstanding potential for the biocatalytic bottom-up synthesis of β-glucans as major bioactive compounds. Their preference for sugar phosphates (rather than nucleotide sugars) as donor substrates further underlines their significance for the carbohydrate industry. Presently, they are classified in the glycoside hydrolase families 94, 149, and 161 (www.cazy.org). Since the discovery of β-1,3-oligoglucan phosphorylase in 1963, several other specificities have been reported that differ in linkage type and/or degree of polymerization. Here, we present an overview of the progress that has been made in our understanding of β-glucan and associated β-glucobiose phosphorylases, with a special focus on their application in the synthesis of carbohydrates and related molecules. Key points: • Discovery, characteristics, and applications of β-glucan phosphorylases. • β-Glucan phosphorylases in the production of functional carbohydrates. [ABSTRACT FROM AUTHOR]
- Published
- 2021
- Full Text
- View/download PDF
7. Expanding the Enzyme Repertoire for Sugar Nucleotide Epimerization: the CDP-Tyvelose 2-Epimerase from Thermodesulfatator atlanticus for Glucose/Mannose Interconversion.
- Author
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Rapp, Christian, van Overtveldt, Stevie, Beerens, Koen, Weber, Hansjörg, Desmet, Tom, and Nidetzky, Bernd
- Subjects
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DEHYDROGENASES , *EPIMERIZATION , *MANNOSE , *NUCLEOTIDE synthesis , *NUCLEAR magnetic resonance , *ENZYMES - Abstract
Epimerization of sugar nucleotides is central to the structural diversification of monosaccharide building blocks for cellular biosynthesis. Epimerase applicability to carbohydrate synthesis can be limited, however, by the high degree of substrate specificity exhibited by most sugar nucleotide epimerases. Here, we discovered a promiscuous type of CDP-tyvelose 2-epimerase (TyvE)-like enzyme that promotes C-2 epimerization in all nucleotide (CDP, UDP, GDP, ADP, and TDP)-activated forms of d-glucose. This new epimerase, originating from Thermodesulfatator atlanticus, is a functional homodimer that contains one tightly bound NAD+/subunit and shows optimum activity at 70°C and pH 9.5. The enzyme exhibits a kcat with CDP-d-glucose of ∼1.0 min−1 (pH 7.5 and 60°C). To characterize the epimerase kinetically and probe its substrate specificity, we developed chemoenzymatic synthesis for CDP-d-mannose, CDP-6-deoxy-d-glucose, CDP-3-deoxy-d-glucose, and CDP-6-deoxy-d-xylo-hexopyranos-4-ulose. Attempts to obtain CDP-d-paratose and CDP-d-tyvelose were not successful. Using high-resolution carbohydrate analytics and in situ nuclear magnetic resonance (NMR) to monitor the enzymatic conversions (60°C and pH 7.5), we show that the CDP-d-mannose/CDP-d-glucose ratio at equilibrium is 0.67 (±0.1), determined from the kinetic Haldane relationship and directly from the reaction. We further show that deoxygenation at sugar C-6 enhances the enzyme activity 5-fold compared to CDP-d-glucose, whereas deoxygenation at C-3 renders the substrate inactive. Phylogenetic analysis places the T. atlanticus epimerase into a distinct subgroup within the sugar nucleotide epimerase family of SDRs (short-chain dehydrogenases/reductases), for which the current study now provides functional context. Collectively, our results expand an emerging toolbox of epimerase-catalyzed reactions for sugar nucleotide synthesis. IMPORTANCE Epimerases of the sugar nucleotide-modifying class of enzymes have attracted considerable interest in carbohydrate (bio)chemistry for the mechanistic challenges and the opportunities for synthesis involved in the reactions catalyzed. The discovery of new epimerases with an expanded scope of sugar nucleotide substrates used is important to promote mechanistic inquiry and can facilitate the development of new enzyme applications. Here, a CDP-tyvelose 2-epimerase-like enzyme from Thermodesulfatator atlanticus is shown to catalyze sugar C-2 epimerization in CDP-glucose and other nucleotide-activated forms of d-glucose. The reactions are new to nature in the context of enzymatic sugar nucleotide modification. The current study explores the substrate scope of the discovered C-2 epimerase and, based on modeling, suggests structure-function relationships that may be important for specificity and catalysis. [ABSTRACT FROM AUTHOR]
- Published
- 2021
- Full Text
- View/download PDF
8. Redirecting substrate regioselectivity using engineered ΔN123-GBD-CD2 branching sucrases for the production of pentasaccharide repeating units of S. flexneri 3a, 4a and 4b haptens.
- Author
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Benkoulouche, Mounir, Ben Imeddourene, Akli, Barel, Louis-Antoine, Le Heiget, Guillaume, Pizzut, Sandra, Kulyk, Hanna, Bellvert, Floriant, Bozonnet, Sophie, Mulard, Laurence A., Remaud-Siméon, Magali, Moulis, Claire, and André, Isabelle
- Subjects
- *
REGIOSELECTIVITY (Chemistry) , *OLIGOSACCHARIDE synthesis , *SHIGELLA flexneri , *HAPTENS , *CARBOHYDRATE synthesis - Abstract
The (chemo-)enzymatic synthesis of oligosaccharides has been hampered by the lack of appropriate enzymatic tools with requisite regio- and stereo-specificities. Engineering of carbohydrate-active enzymes, in particular targeting the enzyme active site, has notably led to catalysts with altered regioselectivity of the glycosylation reaction thereby enabling to extend the repertoire of enzymes for carbohydrate synthesis. Using a collection of 22 mutants of ΔN123-GBD-CD2 branching sucrase, an enzyme from the Glycoside Hydrolase family 70, containing between one and three mutations in the active site, and a lightly protected chemically synthesized tetrasaccharide as an acceptor substrate, we showed that altered glycosylation product specificities could be achieved compared to the parental enzyme. Six mutants were selected for further characterization as they produce higher amounts of two favored pentasaccharides compared to the parental enzyme and/or new products. The produced pentasaccharides were shown to be of high interest as they are precursors of representative haptens of Shigella flexneri serotypes 3a, 4a and 4b. Furthermore, their synthesis was shown to be controlled by the mutations introduced in the active site, driving the glucosylation toward one extremity or the other of the tetrasaccharide acceptor. To identify the molecular determinants involved in the change of ΔN123-GBD-CD2 regioselectivity, extensive molecular dynamics simulations were carried out in combination with in-depth analyses of amino acid residue networks. Our findings help to understand the inter-relationships between the enzyme structure, conformational flexibility and activity. They also provide new insight to further engineer this class of enzymes for the synthesis of carbohydrate components of bacterial haptens. [ABSTRACT FROM AUTHOR]
- Published
- 2021
- Full Text
- View/download PDF
9. An efficient synthesis of 1,6-anhydro-N-acetylmuramic acid from N-acetylglucosamine
- Author
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Matthew B. Calvert, Christoph Mayer, and Alexander Titz
- Subjects
N-acetylmuramic acid ,anhydrosugars ,antibiotic resistance ,bacterial cell wall recycling ,carbohydrate synthesis ,Science ,Organic chemistry ,QD241-441 - Abstract
A novel synthesis of 1,6-anhydro-N-acetylmuramic acid is described, which proceeds in only five steps from the cheap starting material N-acetylglucosamine. This efficient synthesis should enable future studies into the importance of 1,6-anhydromuramic acid in bacterial cell wall recycling processes.
- Published
- 2017
- Full Text
- View/download PDF
10. GmPGL1, a Thiamine Thiazole Synthase, Is Required for the Biosynthesis of Thiamine in Soybean
- Author
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Xingxing Feng, Suxin Yang, Kuanqiang Tang, Yaohua Zhang, Jiantian Leng, Jingjing Ma, Quan Wang, and Xianzhong Feng
- Subjects
soybean ,GmPGL1 ,thiamine thiazole synthase ,carbohydrate synthesis ,amino acid synthesis ,Plant culture ,SB1-1110 - Abstract
Thiamine is an essential cofactor in several enzymatic reactions for all living organisms. Animals cannot synthesize thiamine and depend on their diet. Enhancing the content of thiamine is one of the most important goals of plant breeding to solve the thiamine deficiency associated with the low-thiamin staple crops. In this study, a Glycinemaxpalegreenleaf 1 (Gmpgl1) mutant was isolated from the EMS mutagenized population of soybean cultivar, Williams 82. Map-based cloning of the GmPGL1 locus revealed a single nucleotide deletion at the 292th nucleotide residue of the first exon of Glyma.10g251500 gene in Gmpgl1 mutant plant, encoding a thiamine thiazole synthase. Total thiamine contents decreased in both seedlings and seeds of the Gmpgl1 mutant. Exogenous application of thiazole restored the pale green leaf phenotype of the mutant. The deficiency of thiamine in Gmpgl1 mutant led to reduced activities of the pyruvate dehydrogenase (PDH) and pyruvate decarboxylase (PDC), and decreased contents of six amino acids as compared to that in the wild type plants. These results revealed that GmPGL1 played an essential role in thiamine thiazole biosynthesis.
- Published
- 2019
- Full Text
- View/download PDF
11. GmPGL1 , a Thiamine Thiazole Synthase, Is Required for the Biosynthesis of Thiamine in Soybean.
- Author
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Feng, Xingxing, Yang, Suxin, Tang, Kuanqiang, Zhang, Yaohua, Leng, Jiantian, Ma, Jingjing, Wang, Quan, and Feng, Xianzhong
- Subjects
VITAMIN B1 ,BIOSYNTHESIS ,PLANT breeding ,WILD plants - Abstract
Thiamine is an essential cofactor in several enzymatic reactions for all living organisms. Animals cannot synthesize thiamine and depend on their diet. Enhancing the content of thiamine is one of the most important goals of plant breeding to solve the thiamine deficiency associated with the low-thiamin staple crops. In this study, a Glycine max pale green leaf 1 (Gmpgl1) mutant was isolated from the EMS mutagenized population of soybean cultivar, Williams 82. Map-based cloning of the GmPGL1 locus revealed a single nucleotide deletion at the 292th nucleotide residue of the first exon of Glyma.10g251500 gene in Gmpgl1 mutant plant, encoding a thiamine thiazole synthase. Total thiamine contents decreased in both seedlings and seeds of the Gmpgl1 mutant. Exogenous application of thiazole restored the pale green leaf phenotype of the mutant. The deficiency of thiamine in Gmpgl1 mutant led to reduced activities of the pyruvate dehydrogenase (PDH) and pyruvate decarboxylase (PDC), and decreased contents of six amino acids as compared to that in the wild type plants. These results revealed that GmPGL1 played an essential role in thiamine thiazole biosynthesis. [ABSTRACT FROM AUTHOR]
- Published
- 2019
- Full Text
- View/download PDF
12. A molecular overview of the primary dystroglycanopathies.
- Author
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Brancaccio, Andrea
- Subjects
MUSCULAR dystrophy ,INTEGRINS ,CARBOHYDRATE synthesis ,BASAL lamina ,SKELETAL muscle ,DYSTROGLYCAN - Abstract
Dystroglycan is a major non‐integrin adhesion complex that connects the cytoskeleton to the surrounding basement membranes, thus providing stability to skeletal muscle. In Vertebrates, hypoglycosylation of α‐dystroglycan has been strongly linked to muscular dystrophy phenotypes, some of which also show variable degrees of cognitive impairments, collectively termed dystroglycanopathies. Only a small number of mutations in the dystroglycan gene, leading to the so called primary dystroglycanopathies, has been described so far, as opposed to the ever‐growing number of identified secondary or tertiary dystroglycanopathies (caused by genetic abnormalities in glycosyltransferases or in enzymes involved in the synthesis of the carbohydrate building blocks). The few mutations found within the autonomous N‐terminal domain of α‐dystroglycan seem to destabilise it to different degrees, without influencing the overall folding and targeting of the dystroglycan complex. On the contrary other mutations, some located at the α/β interface of the dystroglycan complex, seem to be able to interfere with its maturation, thus compromising its stability and eventually leading to the intracellular engulfment and/or partial or even total degradation of the dystroglycan uncleaved precursor. [ABSTRACT FROM AUTHOR]
- Published
- 2019
- Full Text
- View/download PDF
13. Chemical Synthesis and Immunological Evaluation of Fragments of the Multiantennary Group-Specific Polysaccharide of Group B Streptococcus
- Author
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Zhen Wang, Jacopo Enotarpi, Giada Buffi, Alfredo Pezzicoli, Christoph J. Gstöttner, Simone Nicolardi, Evita Balducci, Monica Fabbrini, Maria Rosaria Romano, Gijsbert A. van der Marel, Linda del Bino, Roberto Adamo, and Jeroen D. C. Codée
- Subjects
carbohydrate synthesis ,glycosylation ,phosphoramidites ,synthetic vaccines ,glycoconjugates - Abstract
Group B Streptococcus (GBS) is a Gram-positive bacterium and the most common cause of neonatal blood and brain infections. At least 10 different serotypes exist, that are characterized by their different capsular polysaccharides. The Group B carbohydrate (GBC) is shared by all serotypes and therefore attractive be used in a glycoconjugate vaccine. The GBC is a highly complex multiantennary structure, composed of rhamnose rich oligosaccharides interspaced with glucitol phosphates. We here report the development of a convergent approach to assemble a pentamer, octamer, and tridecamer fragment of the termini of the antennae. Phosphoramidite chemistry was used to fuse the pentamer and octamer fragments to deliver the 13-mer GBC oligosaccharide. Nuclear magnetic resonance spectroscopy of the generated fragments confirmed the structures of the naturally occurring polysaccharide. The fragments were used to generate model glycoconjugate vaccine by coupling with CRM197. Immunization of mice delivered sera that was shown to be capable of recognizing different GBS strains. The antibodies raised using the 13-mer conjugate were shown to recognize the bacteria best and the serum raised against this GBC fragment-mediated opsonophagocytic killing best, but in a capsule dependent manner. Overall, the GBC 13-mer was identified to be a highly promising antigen for incorporation into future (multicomponent) anti-GBS vaccines.
- Published
- 2022
- Full Text
- View/download PDF
14. RNASeq analysis of drought-stressed guayule reveals the role of gene transcription for modulating rubber, resin, and carbohydrate synthesis
- Author
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Daniel C. Ilut, Grisel Ponciano, Yong Gu, Colleen M. McMahan, Naxin Huo, and Chen Dong
- Subjects
Parthenium argentatum ,Molecular biology ,Science ,Carbohydrate synthesis ,Carbohydrates ,Asteraceae ,Article ,Terpene ,Transcriptome ,chemistry.chemical_compound ,Fructan ,Biosynthesis ,Natural rubber ,Gene Expression Regulation, Plant ,Botany ,RNA-Seq ,Gene ,Plant Proteins ,Multidisciplinary ,biology ,fungi ,technology, industry, and agriculture ,food and beverages ,biology.organism_classification ,Adaptation, Physiological ,Droughts ,Computational biology and bioinformatics ,chemistry ,visual_art ,visual_art.visual_art_medium ,Medicine ,Rubber ,Plant sciences ,Resins, Plant ,Biotechnology - Abstract
The drought-adapted shrub guayule (Parthenium argentatum) produces rubber, a natural product of major commercial importance, and two co-products with potential industrial use: terpene resin and the carbohydrate fructan. The rubber content of guayule plants subjected to water stress is higher compared to that of well-irrigated plants, a fact consistently reported in guayule field evaluations. To better understand how drought influences rubber biosynthesis at the molecular level, a comprehensive transcriptome database was built from drought-stressed guayule stem tissues using de novo RNA-seq and genome-guided assembly, followed by annotation and expression analysis. Despite having higher rubber content, most rubber biosynthesis related genes were down-regulated in drought-stressed guayule, compared to well-irrigated plants, suggesting post-transcriptional effects may regulate drought-induced rubber accumulation. On the other hand, terpene resin biosynthesis genes were unevenly affected by water stress, implying unique environmental influences over transcriptional control of different terpene compounds or classes. Finally, drought induced expression of fructan catabolism genes in guayule and significantly suppressed these fructan biosynthesis genes. It appears then, that in guayule cultivation, irrigation levels might be calibrated in such a regime to enable tunable accumulation of rubber, resin and fructan.
- Published
- 2021
15. Mechanism of [CO2] Enrichment Alleviated Drought Stress in the Roots of Cucumber Seedlings Revealed via Proteomic and Biochemical Analysis
- Author
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Yiman Li, Wendong Zhang, Dalong Zhang, Yinjian Zheng, Yaliang Xu, Binbin Liu, and Qingming Li
- Subjects
Inorganic Chemistry ,cucumber roots ,CO2 enrichment ,drought stress ,TMT-based quantitative proteomic ,carbohydrate synthesis ,amino acid metabolism ,Organic Chemistry ,General Medicine ,Physical and Theoretical Chemistry ,Molecular Biology ,Spectroscopy ,Catalysis ,Computer Science Applications - Abstract
Cucumber is one of the most widely cultivated greenhouse vegetables, and its quality and yield are threatened by drought stress. Studies have shown that carbon dioxide concentration ([CO2]) enrichment can alleviate drought stress in cucumber seedlings; however the mechanism of this [CO2] enrichment effect on root drought stress is not clear. In this study, the effects of different drought stresses (simulated with 0, 5% and 10% PEG 6000, i.e., no, moderate, and severe drought stress) and [CO2] (400 μmol·mol−1 and 800 ± 40 μmol·mol−1) on the cucumber seedling root proteome were analyzed using the tandem mass tag (TMT) quantitative proteomics method. The results showed that after [CO2] enrichment, 346 differentially accumulating proteins (DAPs) were found only under moderate drought stress, 27 DAPs only under severe drought stress, and 34 DAPs under both moderate and severe drought stress. [CO2] enrichment promoted energy metabolism, amino acid metabolism, and secondary metabolism, induced the expression of proteins related to root cell wall and cytoskeleton metabolism, effectively maintained the balance of protein processing and degradation, and enhanced the cell wall regulation ability. However, the extent to which [CO2] enrichment alleviated drought stress in cucumber seedling roots was limited under severe drought stress, which may be due to excessive damage to the seedlings.
- Published
- 2022
- Full Text
- View/download PDF
16. Transcriptomic profiling of developing fiber in levant cotton (<italic>Gossypium herbaceum</italic> L.).
- Author
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Parekh, Mithil J., Kumar, Sushil, Fougat, Ranbir S., Zala, Harshvardhan N., and Pandit, Ramesh J.
- Subjects
- *
COTTON , *GENE expression , *DIPLOIDY , *PYROSEQUENCING , *CARBOHYDRATE synthesis - Abstract
Cotton (
Gossypium spp.) is an imperative economic crop of the globe due to its natural textile fiber. Molecular mechanisms of fiber development have been greatly revealed in allotetraploid cotton but remained unexplored inGossypium herbaceum .G. herbaceum can withstand the rigors of nature like drought and pests but produce coarse lint. This undesirable characteristic strongly needs the knowledge of fiber development at molecular basis. The present study reported the transcriptome sequence of the developing fiber ofG. herbaceum on pyrosequencing and its analysis. About 1.38 million raw and 1.12 million quality trimmed reads were obtained followed by de novo assembly-generated 20,125 unigenes containing 14,882 coding sequences (CDs). BLASTx-based test of homology indicated that A1-derived transcripts shared a high similarity withGossypium arboreum (A2). Functional annotation of the CDs using the UniProt categorized them into biological processes, cellular components, and molecular function, COG classification showed that a large number of CDs have significant homology in COG database (6215 CDs), and mapping of CDs with Kyoto Encyclopedia of Genes and Genomes (KEGG) database generated 200 pathways ultimately showing predominant engagement in the fiber development process. Transcription factors were predicted by comparison with Plant Transcription Factor Database, and their differential expression between stages exposed their important regulatory role in fiber development. Differential expression analysis based on reads per kilobase of transcript per million mapped reads (RPKM) value revealed activities of specific gene related to carbohydrate and lipid synthesis, carbon metabolism, energy metabolism, signal transduction, etc., at four stages of fiber development, and was validated by qPCR. Overall, this study will help as a valuable foundation for diploid cotton fiber improvement. [ABSTRACT FROM AUTHOR]- Published
- 2018
- Full Text
- View/download PDF
17. The unique evolution of the carbohydrate‐binding module CBM20 in laforin.
- Author
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Kuchtová, Andrea, Gentry, Matthew S., and Janeček, Štefan
- Subjects
- *
CARBOHYDRATE synthesis , *MOLECULAR evolution , *PHYSIOLOGICAL effects of carbohydrates , *PHOSPHATASE genetics , *PHOSPHATASE regulation - Abstract
Laforin catalyses glycogen dephosphorylation. Mutations in its gene result in Lafora disease, a fatal progressive myoclonus epilepsy, the hallmark being water‐insoluble, hyperphosphorylated carbohydrate inclusions called Lafora bodies. Human laforin consists of an N‐terminal carbohydrate‐binding module (CBM) from family CBM20 and a C‐terminal dual‐specificity phosphatase domain. Laforin is conserved in all vertebrates, some basal metazoans and a small group of protozoans. The present
in silico study defines the evolutionary relationships among the CBM20s of laforin with an emphasis on newly identified laforin orthologues. The study reveals putative laforin orthologues inTrichinella , a parasitic nematode, and identifies two sequence inserts in the CBM20 of laforin from parasitic coccidia. Finally, we identify that the putative laforin orthologues from some protozoa and algae possess more than one CBM20. [ABSTRACT FROM AUTHOR]- Published
- 2018
- Full Text
- View/download PDF
18. Identification and characterization of the Streptococcus pneumoniae type 3 capsule-specific glycoside hydrolase of Paenibacillus species 32352.
- Author
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Middleton, Dustin R, Zhang, Xing, Wantuch, Paeton L, Ozdilek, Ahmet, Liu, Xinyue, LoPilato, Rachel, Gangasani, Nikhil, Bridger, Robert, Wells, Lance, and Linhardt, Robert J
- Subjects
- *
STREPTOCOCCUS pneumoniae , *GLYCOSIDE synthesis , *CARBOHYDRATE synthesis , *GLYCOSIDASES , *NUCLEOTIDE sequencing , *DIAGNOSIS - Abstract
Bacillus circulans Jordan 32352 was isolated from decaying organic matter in the New Jersey soil in the early 1930s. This soil-dwelling bacterium produced an enzyme capable of degrading the type 3 capsular polysaccharide (Pn3P) of Streptococcus pneumoniae (Spn). Early reports of this enzyme, Pn3Pase, demonstrated its inducibility by, and specificity for Pn3P. We set out to identify and clone this enzyme for its recombinant expression and characterization. We first sequenced the genome of this bacterial species, and reclassified the Pn3Pase producing bacterium as Paenibacillus species 32352. We identified the putative protein of Pn3Pase through mass spectrometry-based proteomics and cloned the gene for recombinant expression. We then characterized the oligosaccharide products generated upon the enzymatic depolymerization of Pn3P. Sequence analysis suggests that this glycoside hydrolase belongs to a new carbohydrate-active enzyme GH family. To our knowledge, this is the only enzyme to demonstrate Pn3P depolymerization activity. [ABSTRACT FROM AUTHOR]
- Published
- 2018
- Full Text
- View/download PDF
19. β-Glucan phosphorylases in carbohydrate synthesis
- Author
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Jorick Franceus, Tom Desmet, Marc De Doncker, Zorica Ubiparip, and Koen Beerens
- Subjects
β-Glucans ,0106 biological sciences ,Cellodextrin phosphorylase ,CELLVIBRIO-GILVUS ,beta-Glucans ,Glycoside Hydrolases ,Phosphorylases ,Carbohydrate synthesis ,macromolecular substances ,Nucleotide sugar ,01 natural sciences ,Applied Microbiology and Biotechnology ,Laminaribiose phosphorylase ,03 medical and health sciences ,Glycogen phosphorylase ,chemistry.chemical_compound ,010608 biotechnology ,Humans ,CELLODEXTRIN PHOSPHORYLASE ,beta-Glucan phosphorylases ,Glycoside hydrolase ,β-Glucan phosphorylases ,ACCEPTOR SPECIFICITY ,030304 developmental biology ,Glucan ,chemistry.chemical_classification ,0303 health sciences ,PURIFICATION ,Sugar phosphates ,IN-VITRO ,General Medicine ,Mini-Review ,ENZYMATIC GLYCOSYLATION ,CLOSTRIDIUM-THERMOCELLUM CELLOBIOSE ,Chemistry ,LAMINARIBIOSE PHOSPHORYLASE ,Biochemistry ,chemistry ,Biocatalysis ,Carbohydrate Metabolism ,EUGLENA-GRACILIS ,CRYSTALLIZATION ,Biotechnology - Abstract
Abstract β-Glucan phosphorylases are carbohydrate-active enzymes that catalyze the reversible degradation of β-linked glucose polymers, with outstanding potential for the biocatalytic bottom-up synthesis of β-glucans as major bioactive compounds. Their preference for sugar phosphates (rather than nucleotide sugars) as donor substrates further underlines their significance for the carbohydrate industry. Presently, they are classified in the glycoside hydrolase families 94, 149, and 161 (www.cazy.org). Since the discovery of β-1,3-oligoglucan phosphorylase in 1963, several other specificities have been reported that differ in linkage type and/or degree of polymerization. Here, we present an overview of the progress that has been made in our understanding of β-glucan and associated β-glucobiose phosphorylases, with a special focus on their application in the synthesis of carbohydrates and related molecules. Key points • Discovery, characteristics, and applications of β-glucan phosphorylases. • β-Glucan phosphorylases in the production of functional carbohydrates.
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- 2021
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20. Probing the determinants of the transglycosylation/hydrolysis partition in a retaining α-l-arabinofuranosidase
- Author
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Leila Lo Leggio, Tobias Tandrup, Michael J. O’Donohue, Régis Fauré, Jens-Christian N. Poulsen, Sophie Barbe, Bastien Bissaro, Jiao Zhao, Isabelle André, Claire Dumon, Toulouse Biotechnology Institute (TBI), Institut National des Sciences Appliquées - Toulouse (INSA Toulouse), Institut National des Sciences Appliquées (INSA)-Institut National des Sciences Appliquées (INSA)-Centre National de la Recherche Scientifique (CNRS)-Institut National de Recherche pour l’Agriculture, l’Alimentation et l’Environnement (INRAE), Department of Chemistry [Copenhagen], Faculty of Science [Copenhagen], University of Copenhagen = Københavns Universitet (KU)-University of Copenhagen = Københavns Universitet (KU), Institut National des Sciences Appliquées (INSA)-Université de Toulouse (UT)-Institut National des Sciences Appliquées (INSA)-Université de Toulouse (UT)-Centre National de la Recherche Scientifique (CNRS)-Institut National de Recherche pour l’Agriculture, l’Alimentation et l’Environnement (INRAE), and University of Copenhagen = Københavns Universitet (UCPH)-University of Copenhagen = Københavns Universitet (UCPH)
- Subjects
Models, Molecular ,0106 biological sciences ,Glycosylation ,[SDV.BIO]Life Sciences [q-bio]/Biotechnology ,Glycoside Hydrolases ,Stereochemistry ,Mutant ,Carbohydrate synthesis ,Bioengineering ,Context (language use) ,Crystallography, X-Ray ,01 natural sciences ,03 medical and health sciences ,Hydrolysis ,Residue (chemistry) ,010608 biotechnology ,Molecular interactions ,Glycoside hydrolase ,[SDV.BBM.BC]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Biochemistry [q-bio.BM] ,Molecular Biology ,030304 developmental biology ,0303 health sciences ,biology ,Chemistry ,Engineered transglycosylases ,Active site ,General Medicine ,Acceptor ,Mutation ,Biocatalysis ,biology.protein ,Flexibility ,Biotechnology - Abstract
International audience; The use of retaining glycoside hydrolases as synthetic tools for glycochemistry is highly topical and the focus of considerable research. However, due to the incomplete identification of the molecular determinants of the transglycosylation/hydrolysis partition (t/h), rational engineering of retaining glycoside hydrolases to create transglycosylases remains challenging. Therefore, to understand better the factors that underpin transglycosylation in a GH51 retaining α-l-arabinofuranosidase from Thermobacillus xylanilyticus, the investigation of this enzyme’s active site was pursued. Specifically, the properties of two mutants, F26L and L352M, located in the vicinity of the active site are described, using kinetic and 3D structural analyses and molecular dynamics simulations. The results reveal that the presence of L352M in the context of a triple mutant (also containing R69H and N216W) generates changes both in the donor and acceptor subsites, the latter being the result of a domino-like effect. Overall, the mutant R69H-N216W-L352M displays excellent transglycosylation activity (70 % yield, 78 % transfer rate and reduced secondary hydrolysis of the product). In the course of this study, the central role played by the conserved R69 residue was also reaffirmed. The mutation R69H affects both the catalytic nucleophile and the acid/base, including their flexibility, and has a determinant effect on the t/h partition. Finally, the results reveal that increased loop flexibility in the acceptor subsites creates new interactions with the acceptor, in particular with a hydrophobic binding platform composed of N216W, W248 and W302.
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- 2021
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21. Ten years of CAZypedia: a living encyclopedia of carbohydrate-active enzymes.
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CARBOHYDRATE synthesis , *BIOINFORMATICS , *GLYCOMICS , *GLYCOSYLTRANSFERASES , *POLYSACCHARIDES - Abstract
CAZypedia was initiated in 2007 to create a comprehensive, living encyclopedia of the carbohydrateactive enzymes (CAZymes) and associated carbohydrate-binding modules involved in the synthesis, modification and degradation of complex carbohydrates. CAZypedia is closely connected with the actively curated CAZy database, which provides a sequence-based foundation for the biochemical, mechanistic and structural characterization of these diverse proteins. Now celebrating its 10th anniversary online, CAZypedia is a successful example of dynamic, community-driven and expert-based biocuration. [ABSTRACT FROM AUTHOR]
- Published
- 2018
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22. Binding characteristics of galectin-3 fusion proteins.
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Böcker, Sophia and Elling, Lothar
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- *
GALECTINS , *YELLOW fluorescent protein , *SURFACE plasmon resonance , *ENZYME-linked immunosorbent assay , *CARBOHYDRATE synthesis - Abstract
Galectin-3 modulates cell adhesion and signaling events by specific binding and cross-linking galactoside containing carbohydrate ligands. Proteolytic cleavage by metalloproteinases yields in vivo N-terminally truncated galectin-3 still bearing the carbohydrate recognition domain. Truncated galectin-3 has been demonstrated to act in vivo as a negative inhibitor of galectin-3 due to higher affinity for carbohydrate ligands. We here present our studies on a series of 12 human galectin-3 protein constructs. Truncated galectin-3 (Δ1-62 and Δ1-116) and fusions with SNAP-tag and/or yellow fluorescent protein (YFP) display altered binding efficiencies (ratio of maximum binding signal and apparent affinity constant Kd) to asialofetuin (ASF) in solid-phase enzyme-linked immunosorbent assay (ELISA) and surface plasmon resonance (SPR) binding assays. Galectin-3(Δ1-62) and full-length (native) galectin-3 have highest affinity to ASF in ELISA and SPR experiments, respectively, whereas galectin-3(Δ1-116) shows only weak binding. We demonstrate here for the first time that SNAP-tag and YFP fusions of galectin-3 and truncated galectin-3 proteins improve binding efficiencies to ASF. SNAP-tagged galectin-3, galectin-3(Δ1-62) and galectin-3(Δ1-116) are found with significant (3- to 6-fold) higher binding efficiencies in SPR when compared with native galectin-3. Fusion of truncated galectin-3 with YFP renders binding properties similar to native galectin-3, whereas in combination with SNAP-tag improved binding characteristics are obtained. Our results emphasize the importance of the N-terminal domain of human galectin-3 for ligand binding. Most importantly, in combination with fusion proteins suitable for the design of diagnostic and therapeutic tools binding properties can be beneficially tuned. The resulting novel protein tools may be advantageous for potential galectin-3 directed applications in tumor diagnostics and therapy. [ABSTRACT FROM AUTHOR]
- Published
- 2017
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23. Efficient synthesis of α-galactosyl oligosaccharides using a mutant Bacteroides thetaiotaomicron retaining α-galactosidase ( Bt GH97b).
- Author
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Okuyama, Masayuki, Matsunaga, Kana, Watanabe, Ken‐ichi, Yamashita, Keitaro, Tagami, Takayoshi, Kikuchi, Asako, Ma, Min, Klahan, Patcharapa, Mori, Haruhide, Yao, Min, and Kimura, Atsuo
- Subjects
- *
OLIGOSACCHARIDE synthesis , *BACTEROIDES thetaiotaomicron , *NUCLEOPHILES , *GLYCOSIDASES , *HYDROGEN bonding , *LIGAND binding (Biochemistry) , *LINKAGE (Genetics) - Abstract
The preparation of a glycosynthase, a catalytic nucleophile mutant of a glycosidase, is a well-established strategy for the effective synthesis of glycosidic linkages. However, glycosynthases derived from α-glycosidases can give poor yields of desired products because they require generally unstable β-glycosyl fluoride donors. Here, we investigate a transglycosylation catalyzed by a catalytic nucleophile mutant derived from a glycoside hydrolase family ( GH) 97 α-galactosidase, using more stable β-galactosyl azide and α-galactosyl fluoride donors. The mutant enzyme catalyzes the glycosynthase reaction using β-galactosyl azide and α-galactosyl transfer from α-galactosyl fluoride with assistance of external anions. Formate was more effective at restoring transfer activity than azide. Kinetic analysis suggests that poor transglycosylation in the presence of the azide is because of low activity of the ternary complex between enzyme, β-galactosyl azide and acceptor. A three-dimensional structure of the mutant enzyme in complex with the transglycosylation product, β-lactosyl α- d-galactoside, was solved to elucidate the ligand-binding aspects of the α-galactosidase. Subtle differences at the β→α loops 1, 2 and 3 of the catalytic TIM barrel of the α-galactosidase from those of a homologous GH97 α-glucoside hydrolase seem to be involved in substrate recognitions. In particular, the Trp residues in β→α loop 1 have separate roles. Trp312 of the α-galactosidase appears to exclude the equatorial hydroxy group at C4 of glucosides, whereas the corresponding Trp residue in the α-glucoside hydrolase makes a hydrogen bond with this hydroxy group. The mechanism of α-galactoside recognition is conserved among GH27, 31, 36 and 97 α-galactosidases. Database The atomic coordinates (code: ) have been deposited in the Protein Data Bank. [ABSTRACT FROM AUTHOR]
- Published
- 2017
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24. Interactive effects of light quality and culturing temperature on algal cell size, biomass doubling time, protein content, and carbohydrate content
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Tyler W. Johannes, Daniel W. Crunkleton, Jacob Manuel, Shelyn Slavens, and Xiangpeng Li
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0303 health sciences ,Cell division ,030306 microbiology ,Chemistry ,Carbohydrates ,Temperature ,Carbohydrate synthesis ,Biomass ,General Medicine ,Carbohydrate ,Applied Microbiology and Biotechnology ,03 medical and health sciences ,Light intensity ,Nutrient ,Carbohydrate Metabolism ,Doubling time ,Food science ,Growth rate ,Cell Size ,030304 developmental biology ,Biotechnology - Abstract
Light management strategy can be used to improve algal biomass and nutrient production. However, the response of algal metabolism to different light qualities, especially their interaction with other environmental factors, is not well understood. This study focuses on the interactive effects of light quality and culturing temperature on algal protein content and carbohydrate content of C. reinhardtii. Three LED light sources (blue light, red-orange light, and white-yellow light) were applied to grow algae in batch cultures with a light intensity of 105 μmol/m2s under the temperatures of 24 °C to 32 °C. The protein and carbohydrate content were measured in both the late exponential growth phase and the late stationary growth phase. The results revealed that there was an interactive effect of light quality and culturing temperature on the protein and carbohydrate content. The combined conditions of blue light and a temperature of 24 °C or 28 °C, which induced a larger algal cell size with a prolonged cell cycle and a low division rate, resulted in the highest protein content; the protein mass fraction and concentration were 32% and 52% higher than that under white-yellow light at 32 °C. The combined conditions of red-orange light and a temperature of 24 °C, which promoted both the cell division and size growth, enhanced the carbohydrate content; the carbohydrate mass fraction and concentration were 161% and 155% higher than that under white-yellow light at 24 °C. When there was temperature stress (32 °C) or nutrient stress, the effect of light quality reduced, and the difference of protein and carbohydrate content among the three light qualities decreased. KEY POINTS: • Studied light quality-temperature interactive effect on protein, carbohydrate synthesis. • Protein content was high under low cell division rate. • Carbohydrate content was high under high cell division and cell size growth rate.
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- 2021
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25. Glycosyl disulfides: importance, synthesis and application to chemical and biological systems
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Robert A. Falconer and Goreti Ribeiro Morais
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Anomer ,010405 organic chemistry ,Carbohydrate chemistry ,Glycobiology ,Organic Chemistry ,Disulfide bond ,Carbohydrate synthesis ,Chemistry Techniques, Synthetic ,010402 general chemistry ,01 natural sciences ,Biochemistry ,Combinatorial chemistry ,0104 chemical sciences ,carbohydrates (lipids) ,chemistry.chemical_compound ,chemistry ,Functional group ,Animals ,Humans ,Molecule ,Glycosyl ,Disulfides ,Glycosides ,Physical and Theoretical Chemistry ,Glycomics - Abstract
The disulfide bond plays an important role in the formation and stabilisation of higher order structures of peptides and proteins, while in recent years interest in this functional group has been extended to carbohydrate chemistry. Rarely found in nature, glycosyl disulfides have attracted significant attention as glycomimetics, with wide biological applications including lectin binding, as key components of dynamic libraries to study carbohydrate structures, the study of metabolic and enzymatic studies, and even as potential drug molecules. This interest has been accompanied and fuelled by the continuous development of new methods to construct the disulfide bond at the anomeric centre. Glycosyl disulfides have also been exploited as versatile intermediates in carbohydrate synthesis, particularly as glycosyl donors. This review focuses on the importance of the disulfide bond in glycobiology and in chemistry, evaluating the different methods available to synthesise glycosyl disulfides. Furthermore, we review the role of glycosyl disulfides as intermediates and/or glycosyl donors for the synthesis of neoglycoproteins and oligosaccharides, before finally considering examples of how this important class of carbohydrates have made an impact in biological and therapeutic contexts.
- Published
- 2021
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26. Characterization of Mannosyl Dioxanium Ions in Solution Using Chemical Exchange Saturation Transfer NMR Spectroscopy
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Sam J. Moons, Frank F J de Kleijne, Paul B. White, Thomas J. Boltje, and Hidde Elferink
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chemistry.chemical_classification ,Glycosylation ,Metal ions in aqueous solution ,Kinetics ,Reactive intermediate ,Carbohydrate synthesis ,Glycosidic bond ,General Medicine ,Synthetic Organic Chemistry ,General Chemistry ,Nuclear magnetic resonance spectroscopy ,Catalysis ,carbohydrates (lipids) ,chemistry.chemical_compound ,chemistry ,Computational chemistry ,lipids (amino acids, peptides, and proteins) ,Acyl group - Abstract
The stereoselective introduction of the glycosidic bond remains one of the main challenges in carbohydrate synthesis. Characterizing the reactive intermediates of this reaction is key to develop stereoselective glycosylation reactions. Herein we report the characterization of low-populated, rapidly equilibrating, mannosyl dioxanium ions that arise from participation of a C-3 acyl group using chemical exchange saturation transfer (CEST) NMR spectroscopy. Dioxanium ion structure and equilibration kinetics were measured under relevant glycosylation conditions and highly α -selective couplings were observed suggesting glycosylation took place via this elusive intermediate.
- Published
- 2022
27. Synthesis of a homologous series of galactofuranose-containing mycobacterial arabinogalactan fragments.
- Author
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Joe, Maju and Lowary, Todd L.
- Subjects
- *
FURANOSIDES , *CARBOHYDRATE synthesis , *ARABINOGALACTAN , *MYCOBACTERIA , *PEPTIDOGLYCANS , *BIOSYNTHESIS - Abstract
Mycobacteria, including the human pathogen Mycobacterium tuberculosis, the causative agent of tuberculosis, produce a complex cell wall structure made of carbohydrates and lipids. The major structural element of the mycobacterial cell wall is a glycoconjugate called the mycolic acid - arabinogalactan - peptidoglycan (mAGP) complex. Inhibition of mAGP biosynthesis is a proven strategy for developing anti-mycobacterial drugs, and thus, understanding the pathways and enzymes involved in the assembly of this molecule is of interest. In this paper, we describe the chemical synthesis of a panel of nine oligosaccharide fragments ( 4- 12) of the galactan domain of the mAGP complex designed as biosynthetic probes. These structures, ranging in size from a hexasaccharide to a tetradecasaccharide, are potential substrates for two biosynthetic enzymes, GlfT2 and AftA, and represent the largest mycobacterial galactan fragments synthesized to date. The route developed was iterative and provided multimilligram quantities of the target molecules 4- 12 in good overall yield. [ABSTRACT FROM AUTHOR]
- Published
- 2016
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28. SpyB, a Small Heme-Binding Protein, Affects the Composition of the Cell Wall in Streptococcus pyogenes.
- Author
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Edgar, Rebecca J., Jing Chen, Kant, Sashi, Rechkina, Elena, Rush, Jeffrey S., Forsberg, Lennart S., Jaehrig, Bernhard, Azadi, Parastoo, Tchesnokova, Veronika, Sokurenko, Evgeni V., Haining Zhu, Korotkov, Konstantin V., Pancholi, Vijay, and Korotkova, Natalia
- Subjects
STREPTOCOCCUS pyogenes ,CARBOHYDRATE synthesis ,LACTAM derivatives ,GLYCOSIDES ,SACCHARIDES ,DISULFIDES synthesis ,BACTERIAL cell walls - Abstract
Streptococcus pyogenes (Group A Streptococcus or GAS) is a hemolytic human pathogen associated with a wide variety of infections ranging from minor skin and throat infections to life-threatening invasive diseases. The cell wall of GAS consists of peptidoglycan sacculus decorated with a carbohydrate comprising a polyrhamnose backbone with immunodominant N-acetylglucosamine side-chains. All GAS genomes contain the spyBA operon, which encodes a 35-amino-acid membrane protein SpyB, and a membrane-bound C3-like ADP-ribosyltransferase SpyA. In this study, we addressed the function of SpyB in GAS. Phenotypic analysis of a spyB deletion mutant revealed increased bacterial aggregation, and reduced sensitivity to b-lactams of the cephalosporin class and peptidoglycan hydrolase PlyC. Glycosyl composition analysis of cell wall isolated from the spyB mutant suggested an altered carbohydrate structure compared with the wild-type strain. Furthermore, we found that SpyB associates with heme and protoporphyrin IX. Heme binding induces SpyB dimerization, which involves disulfide bond formation between the subunits. Thus, our data suggest the possibility that SpyB activity is regulated by heme. [ABSTRACT FROM AUTHOR]
- Published
- 2016
- Full Text
- View/download PDF
29. Identification of Genes Potentially Associated with the Fertility Instability of S-Type Cytoplasmic Male Sterility in Maize via Bulked Segregant RNA-Seq.
- Author
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Su, Aiguo, Song, Wei, Xing, Jinfeng, Zhao, Yanxin, Zhang, Ruyang, Li, Chunhui, Duan, Minxiao, Luo, Meijie, Shi, Zi, and Zhao, Jiuran
- Subjects
- *
CYTOPLASMIC male sterility , *RNA sequencing , *QUANTITATIVE genetics , *SINGLE nucleotide polymorphisms , *CARBOHYDRATE synthesis ,CORN genetics - Abstract
S-type cytoplasmic male sterility (CMS-S) is the largest group among the three major types of CMS in maize. CMS-S exhibits fertility instability as a partial fertility restoration in a specific nuclear genetic background, which impedes its commercial application in hybrid breeding programs. The fertility instability phenomenon of CMS-S is controlled by several minor quantitative trait locus (QTLs), but not the major nuclear fertility restorer (Rf3). However, the gene mapping of these minor QTLs and the molecular mechanism of the genetic modifications are still unclear. Using completely sterile and partially rescued plants of fertility instable line (FIL)-B, we performed bulk segregant RNA-Seq and identified six potential associated genes in minor effect QTLs contributing to fertility instability. Analyses demonstrate that these potential associated genes may be involved in biological processes, such as floral organ differentiation and development regulation, energy metabolism and carbohydrates biosynthesis, which results in a partial anther exsertion and pollen fertility restoration in the partially rescued plants. The single nucleotide polymorphisms (SNPs) identified in two potential associated genes were validated to be related to the fertility restoration phenotype by KASP marker assays. This novel knowledge contributes to the understanding of the molecular mechanism of the partial fertility restoration of CMS-S in maize and thus helps to guide the breeding programs. [ABSTRACT FROM AUTHOR]
- Published
- 2016
- Full Text
- View/download PDF
30. Cellular Composition Changes and Nitrogen Uptake under Extra-Limited Nitrogen Conditions by Thermosynechococcus sp. CL-1 Carbon Biofixation.
- Author
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Chi-Ming, Tseng, Tzu-Hsing, Ko, Hsin-Ta, Hsueh, Hsing-Hui, Chen, Dah-Tong, Ray, Yun-Hwei, Shen, and Hsin, Chu
- Subjects
- *
BACTERIAL cell analysis , *CARBON fixation , *NITROGEN , *THERMOSYNECHOCOCCUS elongatus , *PHOTOSYNTHETIC bacteria , *CARBOHYDRATE synthesis , *SPECTROPHOTOMETERS - Published
- 2016
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31. Biosynthesis and Biological Activity of Carbasugars.
- Author
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Roscales, Silvia and Plumet, Joaquín
- Subjects
- *
BIOSYNTHESIS , *MONOSACCHARIDES , *FUNCTIONAL groups , *PYRANOSES , *CARBOHYDRATE synthesis , *BIOACTIVE compounds - Abstract
The first synthesis of carbasugars, compounds in which the ring oxygen of a monosaccharide had been replaced by a methylene moiety, was described in 1966 by Professor G. E. McCasland’s group. Seven years later, the first true natural carbasugar (5a-carba-R-D-galactopyranose) was isolated from a fermentation broth of Streptomyces sp. MA-4145. In the following decades, the chemistry and biology of carbasugars have been extensively studied. Most of these compounds show interesting biological properties, especially enzymatic inhibitory activities, and, in consequence, an important number of analogues have also been prepared in the search for improved biological activities. The aim of this review is to give coverage on the progress made in two important aspects of these compounds: the elucidation of their biosynthesis and the consideration of their biological properties, including the extensively studied carbapyranoses as well as the much less studied carbafuranoses. [ABSTRACT FROM AUTHOR]
- Published
- 2016
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32. An Integrated Bioinformatics Analysis Reveals Divergent Evolutionary Pattern of Oil Biosynthesis in High- and Low-Oil Plants.
- Author
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Zhang, Li, Wang, Shi-Bo, Li, Qi-Gang, Song, Jian, Hao, Yu-Qi, Zhou, Ling, Zheng, Huan-Quan, Dunwell, Jim M., and Zhang, Yuan-Ming
- Subjects
- *
OILSEEDS , *BIOMASS energy , *OILSEED plants , *FATTY acid synthesis , *CHROMOSOME duplication , *CARBOHYDRATE synthesis - Abstract
Seed oils provide a renewable source of food, biofuel and industrial raw materials that is important for humans. Although many genes and pathways for acyl-lipid metabolism have been identified, little is known about whether there is a specific mechanism for high-oil content in high-oil plants. Based on the distinct differences in seed oil content between four high-oil dicots (20~50%) and three low-oil grasses (<3%), comparative genome, transcriptome and differential expression analyses were used to investigate this mechanism. Among 4,051 dicot-specific soybean genes identified from 252,443 genes in the seven species, 54 genes were shown to directly participate in acyl-lipid metabolism, and 93 genes were found to be associated with acyl-lipid metabolism. Among the 93 dicot-specific genes, 42 and 27 genes, including CBM20-like SBDs and GPT2, participate in carbohydrate degradation and transport, respectively. 40 genes highly up-regulated during seed oil rapid accumulation period are mainly involved in initial fatty acid synthesis, triacylglyceride assembly and oil-body formation, for example, ACCase, PP, DGAT1, PDAT1, OLEs and STEROs, which were also found to be differentially expressed between high- and low-oil soybean accessions. Phylogenetic analysis revealed distinct differences of oleosin in patterns of gene duplication and loss between high-oil dicots and low-oil grasses. In addition, seed-specific GmGRF5, ABI5 and GmTZF4 were predicted to be candidate regulators in seed oil accumulation. This study facilitates future research on lipid biosynthesis and potential genetic improvement of seed oil content. [ABSTRACT FROM AUTHOR]
- Published
- 2016
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33. Template free synthesis of natural carbohydrates functionalised fluorescent silver nanoclusters.
- Author
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Ebrahiminezhad, Alireza, Berenjian, Aydin, and Ghasemi, Younes
- Subjects
- *
CARBOHYDRATE synthesis , *SILVER nanoparticles , *FLUORESCENCE , *GERMAN chamomile , *FUNCTIONAL groups - Abstract
Template-assisted synthesis is one of the most recognised techniques for fabrication of silver nanoclusters (AgNCs). However, this process is time consuming, toxic and expensive. In this study, the authors report a completely novel approach for the green and facile synthesis of AgNCs using Matricaria chamomilla, without any additional template. Fluorescent and colloidally stable AgNCs with average particle size of 2.4 nm were successfully produced. They found that carbohydrates from Matricaria chamomilla act as an ideal template to generate fluorescent AgNCs. Moreover, oxygen-bearing functional groups were validated to be the active groups for anchoring and reducing of Ag+ ions. The novel carbohydrate coating method makes the prepared nanoclusters completely hydrophilic and stable in aqueous matrices. [ABSTRACT FROM AUTHOR]
- Published
- 2016
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34. Structural insights into the carbohydrate-binding ability of an α-(1→2) branching sucrase from glycoside-hydrolase family 70.
- Author
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Brison, Yoann, Malbert, Yannick, Czaplicki, Georges, Mourey, Lionel, Remaud-Simeon, Magali, and Tranier, Samuel
- Subjects
- *
CARBOHYDRATE synthesis , *GLYCOSIDASES , *DEXTRANSUCRASE , *CATALYTIC hydrogenation , *VAN der Waals forces - Abstract
The α-(1→2) branching sucrase ΔN123-GBD-CD2 is an enzyme belonging to the Glycoside-Hydrolase family 70 (GH70) that catalyses the transfer of D-glucosyl units from sucrose to dextrans or glucooligosaccharides via the formation of α-(1→2) glucosidic linkages. The first structures of ΔN123-GBD-CD2 in complex with D-glucose, isomaltosyl or isomaltotriosyl residues were solved. The glucose complex revealed three glucose binding sites in the catalytic gorge and six additional ones at the surface of domains B, IV and V. Soaking with isomaltotriose or glucooligosaccharides led to structures in which isomaltosyl or isomaltriosyl residues were found in glucan binding pockets belonging to domain V. One aromatic residue is systematically identified at the bottom of these pockets in stacking interaction with one glucosyl moiety. The carbohydrate is also maintained by a network of hydrogen bonds and van der Waals interactions. The sequence of these binding pockets is conserved and repeatedly present in the domain V of GH70 glucansucrases known to bind α-glucans. These findings provide the first structural evidence of the molecular interaction occurring between isomaltooligosaccharides and the domain V of the GH70 enzymes. [ABSTRACT FROM AUTHOR]
- Published
- 2016
- Full Text
- View/download PDF
35. Synthesis of new photoactive urethane carbohydrates and their behavior in UV or femtosecond laser-induced two-photon polymerization.
- Author
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Chibac, Andreea L., Buruiana, Tinca, Melinte, Violeta, Mangalagiu, Ionel, Epurescu, George, and Buruiana, Emil C.
- Subjects
- *
CARBOHYDRATE synthesis , *URETHANE , *FEMTOSECOND lasers , *PHOTOPOLYMERIZATION , *METHACRYLATES , *FOURIER transform spectroscopy , *HYDROPHILIC interactions - Abstract
Three methacrylated glycomonomers (CMA-1–3) were synthesized, characterized, and photocrosslinked in hydrophilic conetworks. The UV photobehavior of monomers was evaluated through FTIR spectrometry using Irgacure 819/Irgacure 2959 as photoinitiator, and the results showed that Irgacure 2959 is a better photoinitiator, the conversion degree (DC) varying in the range of 69–89.2% and the maximum rate of polymerization () being ~10 s−1. The addition of poly(ethylene glycol) dimethacrylate (PEG-DMA) or benzophenone macromer (BP-UDMA) as co-monomer improved theand DC (up to 100% after 300 s of irradiation in the latter). Specific analysis (gel fraction, water uptake, morphology) of the photopolymerized networks A1–A3 (CMA-1–3/PEG-DMA) and B1–B3 (CMA-1–3/BP-UDMA) demonstrated that B1–B3 films have interconnected inner pores, with irregular shapes and different sizes. The grid structures created through the two-photon polymerization of B1–B3 formulations in the presence of Irg819 are clearer and have a controlled porosity, of relevance for promising bioapplications. [ABSTRACT FROM PUBLISHER]
- Published
- 2016
- Full Text
- View/download PDF
36. A mild and convenient approach for selective acetonide cleavage involved in carbohydrate synthesis using PPA-SiO2
- Author
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Rahul R. Nikam and Kiran R. Gore
- Subjects
Primary (chemistry) ,010405 organic chemistry ,Stereochemistry ,Chemistry ,Organic Chemistry ,Carbohydrate synthesis ,respiratory system ,Carbohydrate ,010402 general chemistry ,Highly selective ,Cleavage (embryo) ,01 natural sciences ,Biochemistry ,Acetonide ,0104 chemical sciences ,Selective cleavage - Abstract
Here, we report a highly selective, efficient and rapid method for the selective cleavage of primary acetonide using silica supported polyphosphoric acid (PPA-SiO2) for various carbohydrate substrates. Corresponding diols were obtained in good to excellent yields within 30 min using PPA-SiO2. Overall, PPA-SiO2 was found to be a useful catalyst for selective acetonide cleavage in carbohydrate substrates which may expand its utility in organic synthesis.
- Published
- 2020
- Full Text
- View/download PDF
37. Introducing Oxo-Phenylacetyl (OPAc) as a Protecting Group for Carbohydrates
- Author
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Suhail A. Rather, Qazi Naveed Ahmed, Ram A. Vishwakarma, Veeranjaneyulu Gannedi, and Atul Kumar
- Subjects
chemistry.chemical_classification ,010405 organic chemistry ,Organic Chemistry ,Carbohydrate synthesis ,010402 general chemistry ,01 natural sciences ,Combinatorial chemistry ,0104 chemical sciences ,chemistry.chemical_compound ,chemistry ,Reagent ,Monosaccharide ,Glycosyl ,Protecting group - Abstract
A series of oxo-phenylacetyl (OPAc)-protected saccharides, with divergent base sensitivity profiles against benzoyl (Bz) and acetyl (Ac) were synthesized, and KHSO5/AcCl in methanol was identified as an easy, mild, selective, and efficient deprotecting reagent for their removal in the perspective of carbohydrate synthesis. Timely monitoring of AcCl reagent was supportive in both sequential and simultaneous deprotecting of OPAc, Bz, and Ac. The salient feature of our method is the orthogonal stability against different groups, its ease to generate different valuable acceptors using designed monosaccharides, and use of OPAc as a glycosyl donar.
- Published
- 2019
- Full Text
- View/download PDF
38. Tools for understanding the glycosaminoglycan biosynthesis
- Author
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Willén, Daniel and Willén, Daniel
- Abstract
For cells to function correctly within an organism, they need different systems to communicate with each other. One crucial part of cell signaling is the proteoglycans (PGs) and glycosaminoglycans (GAGs), which are macromolecules that bind different signaling molecules and proteins necessary for processes such as growth and proliferation. Therefore, PGs and GAGs are involved in pathological processes like cancer and bacterial or viral infection. A tetrasaccharide linker connects several types of GAGs to the PG core protein, with the first carbohydrate being a xylose. Xylose is, due to its scarcity in mammalian cells, an attractive target for therapeutics. In GAG biosynthesis, the enzyme β4GalT7 galactosylates the xylose. Therefore, this enzyme could be interesting to target using synthetic xylosides that could act as substrates or inhibitors. These xylosides could allow us to understand and control the biosynthesis of GAGs. This thesis is focused on the design and synthesis of modified xylosides and how we can use these as tools to study the formation of GAGs. We decided to alter the carbohydrate itself to investigate if it is possible to design effective substrates or inhibitors for β4GalT7. We also decided to modify the aglycon, the part attached to the carbohydrate, to answer questions related to how cells process GAGs. This work has resulted in several new substances that, that enter cells, and work well as substrates and inhibitors of β4GalT7 to provide answers to how GAGs are structured, how they move within the cell, and how they interact with other biomolecules such as viral proteins. The results of this work pave the way for the use of synthetic xylosides to answer several questions about GAG-related processes and open up the possibility for creating tools for influencing and studying cells' expression of GAGs.
- Published
- 2021
39. PPP2R5C Couples Hepatic Glucose and Lipid Homeostasis.
- Author
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Cheng, Yong-Sheng, Seibert, Oksana, Klöting, Nora, Dietrich, Arne, Straßburger, Katrin, Fernández-Veledo, Sonia, Vendrell, Joan J., Zorzano, Antonio, Blüher, Matthias, Herzig, Stephan, Berriel Diaz, Mauricio, and Teleman, Aurelio A.
- Subjects
- *
LIVER , *HOMEOSTASIS , *LIPIDS , *GLUCOSE , *CARBOHYDRATE synthesis , *MAMMALS - Abstract
In mammals, the liver plays a central role in maintaining carbohydrate and lipid homeostasis by acting both as a major source and a major sink of glucose and lipids. In particular, when dietary carbohydrates are in excess, the liver converts them to lipids via de novo lipogenesis. The molecular checkpoints regulating the balance between carbohydrate and lipid homeostasis, however, are not fully understood. Here we identify PPP2R5C, a regulatory subunit of PP2A, as a novel modulator of liver metabolism in postprandial physiology. Inactivation of PPP2R5C in isolated hepatocytes leads to increased glucose uptake and increased de novo lipogenesis. These phenotypes are reiterated in vivo, where hepatocyte specific PPP2R5C knockdown yields mice with improved systemic glucose tolerance and insulin sensitivity, but elevated circulating triglyceride levels. We show that modulation of PPP2R5C levels leads to alterations in AMPK and SREBP-1 activity. We find that hepatic levels of PPP2R5C are elevated in human diabetic patients, and correlate with obesity and insulin resistance in these subjects. In sum, our data suggest that hepatic PPP2R5C represents an important factor in the functional wiring of energy metabolism and the maintenance of a metabolically healthy state. [ABSTRACT FROM AUTHOR]
- Published
- 2015
- Full Text
- View/download PDF
40. Identification of carbohydrate anomers using ion mobility-mass spectrometry.
- Author
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Hofmann, J., Hahm, H. S., Seeberger, P. H., and Pagel, K.
- Subjects
- *
CARBOHYDRATE analysis , *ANOMERS , *ION mobility spectroscopy , *MASS spectrometry , *STEREOISOMERS , *CARBOHYDRATE synthesis - Abstract
Carbohydrates are ubiquitous biological polymers that are important in a broad range of biological processes. However, owing to their branched structures and the presence of stereogenic centres at each glycosidic linkage between monomers, carbohydrates are harder to characterize than are peptides and oligonucleotides. Methods such as nuclear magnetic resonance spectroscopy can be used to characterize glycosidic linkages, but this technique requires milligram amounts of material and cannot detect small amounts of coexisting isomers. Mass spectrometry, on the other hand, can provide information on carbohydrate composition and connectivity for even small amounts of sample, but it cannot be used to distinguish between stereoisomers. Here, we demonstrate that ion mobility-mass spectrometry-a method that separates molecules according to their mass, charge, size, and shape-can unambiguously identify carbohydrate linkage-isomers and stereoisomers. We analysed six synthetic carbohydrate isomers that differ in composition, connectivity, or configuration. Our data show that coexisting carbohydrate isomers can be identified, and relative concentrations of the minor isomer as low as 0.1 per cent can be detected. In addition, the analysis is rapid, and requires no derivatization and only small amounts of sample. These results indicate that ion mobility-mass spectrometry is an effective tool for the analysis of complex carbohydrates. This method could have an impact on the field of carbohydrate synthesis similar to that of the advent of high-performance liquid chromatography on the field of peptide assembly in the late 1970s. [ABSTRACT FROM AUTHOR]
- Published
- 2015
- Full Text
- View/download PDF
41. Bioactive Carbohydrates and Peptides in Foods: An Overview of Sources, Downstream Processing Steps and Associated Bioactivities.
- Author
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Hayes, Maria and Tiwari, Brijesh K.
- Subjects
- *
PEPTIDE synthesis , *CARBOHYDRATE synthesis , *BIOACTIVE compounds , *MANUFACTURING processes , *REFINED carbohydrates - Abstract
Bioactive peptides and carbohydrates are sourced from a myriad of plant, animal and insects and have huge potential for use as food ingredients and pharmaceuticals. However, downstream processing bottlenecks hinder the potential use of these natural bioactive compounds and add cost to production processes. This review discusses the health benefits and bioactivities associated with peptides and carbohydrates of natural origin and downstream processing methodologies and novel processes which may be used to overcome these. [ABSTRACT FROM AUTHOR]
- Published
- 2015
- Full Text
- View/download PDF
42. An Unusual Carbohydrate Conformation is Evident in Moraxella catarrhalis Oligosaccharides.
- Author
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Frank, Martin, Collins, Patrick M., Peak, Ian R., Grice, I. Darren, and Wilson, Jennifer C.
- Subjects
- *
OLIGOSACCHARIDE structure , *CONFORMATIONAL analysis , *MORAXELLA catarrhalis , *CARBOHYDRATE synthesis , *LIPOOLIGOSACCHARIDES , *INTRAMOLECULAR catalysis , *HYDROPHOBIC interactions - Abstract
Oligosaccharide structures derived from the lipooligosaccharide of M. catarrhalis show that the highly branched glucose-rich inner core of the oligosaccharide has an altered conformation compared to the most truncated tetra-glucose-Kdo lgt1/4Δ oligosaccharide structure. Addition of one residue each to the (1-4) and (1-6) chains to give the lgt2Δ oligosaccharide is the minimum requirement for this conformational change to occur. Extensive molecular modeling and NMR investigations have shown that the (1-3), (1-4), and (1-6) glycosidic linkages from the central α-D-Glcp have significantly altered conformational preferences between the two structures. For the lgt1/4Δ oligosaccharide the (1-3) and (1-4) linkage populates predominantly the syn minimum on the conformational free energy map and for the (1-6) linkage conformational flexibility is observed, which is supported by 1H-NMR T1 measurements. For the lgt2Δ oligosaccharide the unusual “(1-4)anti-ψ(1-6)gg” conformation, which could be confirmed by long-range NOE signals, is a dominant conformation in which the oligosaccharide is very compact with the terminal α-D-GlcNAc residue folding back towards the center of the molecule leading to an extensive intra-molecular hydrophobic interaction between the terminal residues. Comparing effective H-H distances, which were calculated for conformational sub-ensembles, with the NOE distances revealed that typically multiple conformations could be present without significantly violating the measured NOE restraints. For lgt2Δ the presence of more than one conformation is supported by the NOE data. [ABSTRACT FROM AUTHOR]
- Published
- 2015
- Full Text
- View/download PDF
43. Bio-Refining of Carbohydrate-Rich Food Waste for Biofuels.
- Author
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Hoang-Tuong Nguyen Hao, Karthikeyan, Obulisamy Parthiba, and Heimann, Kirsten
- Subjects
- *
CARBOHYDRATE synthesis , *BIOMASS burning , *BIOMASS energy , *FATTY acid methyl esters , *HYDROLYSIS kinetics - Abstract
The global dependence on finite fossil fuel-derived energy is of serious concern given the predicted population increase. Over the past decades, bio-refining of woody biomass has received much attention, but data on food waste refining are sorely lacking, despite annual and global deposition of 1.3 billion tons in landfills. In addition to negative environmental impacts, this represents a squandering of valuable energy, water and nutrient resources. The potential of carbohydrate-rich food waste (CRFW) for biofuel (by Rhodotorulla glutinis fermentation) and biogas production (by calculating theoretical methane yield) was therefore investigated using a novel integrated bio-refinery approach. In this approach, hydrolyzed CRFW from three different conditions was used for Rhodotorulla glutinis cultivation to produce biolipids, whilst residual solids after hydrolysis were characterized for methane recovery potential via anaerobic digestion. Initially, CRFW was hydrolysed using thermal- (Th), chemical- (Ch) and Th-Ch combined hydrolysis (TCh), with the CRFW-leachate serving as a control (Pcon). Excessive foaming led to the loss of TCh cultures, while day-7 biomass yields were similar (3.4-3.6 g dry weight (DW) L-1) for the remaining treatments. Total fatty acid methyl ester (FAME) content of R. glutinis cultivated on CRFW hydrolysates were relatively low (~6.5%) but quality parameters (i.e., cetane number, density, viscosity and higher heating values) of Energies 2015, 8 6351 biomass extracted biodiesel complied with ASTM standards. Despite low theoretical RS-derived methane potential, further research under optimised and scaled conditions will reveal the potential of this approach for the bio-refining of CRFW for energy recovery and value-added co-product production. [ABSTRACT FROM AUTHOR]
- Published
- 2015
- Full Text
- View/download PDF
44. Planteose as a storage carbohydrate required for early stage of germination of Orobanche minor and its metabolism as a possible target for selective control.
- Author
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Takatoshi Wakabayashi, Joseph, Benesh, Shuhei Yasumoto, Tomoyoshi Akashi, Toshio Aoki, Kazuo Harada, Satoru Muranaka, Takeshi Bamba, Eiichiro Fukusaki, Yasutomo Takeuchi, Koichi Yoneyama, Toshiya Muranaka, Yukihiro Sugimoto, and Atsushi Okazawa
- Subjects
- *
BROOMRAPES , *PHYSIOLOGICAL effects of carbohydrates , *CARBOHYDRATE synthesis , *BIOLOGICAL weed control , *METABOLIC regulation - Abstract
Root parasitic weeds in Orobanchaceae cause serious damage to worldwide agriculture. Germination of the parasites requires host-derived germination stimulants, such as strigolactones, as indicators of host roots within reach of the parasite's radicles. This unique germination process was focused on to identify metabolic pathways required for germination, and to design a selective control strategy. A metabolomic analysis of germinating seeds of clover broomrape, Orobanche minor, was conducted to identify its distinctive metabolites. Consequently, a galactosylsucrose trisaccharide, planteose (α-d-galactopyranosyl-(1→6)-β-d-fructofuranosyl-(2→1)-α-d-glucopyranoside), was identified as a metabolite that decreased promptly after reception of the germination stimulant. To investigate the importance of planteose metabolism, the effects of several glycosidase inhibitors were examined, and nojirimycin bisulfite (NJ) was found to alter the sugar metabolism and to selectively inhibit the germination of O. minor. Planteose consumption was similar in NJ-treated seeds and non-treated germinating seeds; however, NJ-treated seeds showed lower consumption of sucrose, a possible intermediate of planteose metabolism, resulting in significantly less glucose and fructose. This inhibitory effect was recovered by adding glucose. These results suggest that planteose is a storage carbohydrate required for early stage of germination of O. minor, and NJ inhibits germination by blocking the supply of essential glucose from planteose and sucrose. Additionally, NJ selectively inhibited radicle elongation of germinated seeds of Orobanchaceae plants (Striga hermonthica and Phtheirospermum japonicum). Thus, NJ will be a promising tool to develop specific herbicides to the parasites, especially broomrapes, and to improve our understanding of the molecular mechanisms of this unique germination. [ABSTRACT FROM AUTHOR]
- Published
- 2015
- Full Text
- View/download PDF
45. A Survey of Recent Synthetic Applications of 2,3-Dideoxy-Hex-2-enopyranosides.
- Author
-
Gomez, Ana M., Lobo, Fernando, Miranda, Silvia, and Cristobal Lopez, J.
- Subjects
- *
PYRANOSIDE , *RING formation (Chemistry) , *CARBOHYDRATE synthesis , *UNSATURATED compounds , *SYNTHETIC products , *EPOXIDATION , *PHASE transitions - Abstract
Unsaturated carbohydrate derivatives are useful intermediates in synthetic transformations leading to a variety of compounds. The aim of this review is to highlight the rich chemistry of Δ-2,3 unsaturated pyranosides, emphasizing the variety of transformations that have been carried out in these substrates during the last decade. [ABSTRACT FROM AUTHOR]
- Published
- 2015
- Full Text
- View/download PDF
46. Finding the C4 sweet spot: cellular compartmentation of carbohydrate metabolism in C4 photosynthesis
- Author
-
Robert T. Furbank and Steven L. Kelly
- Subjects
0106 biological sciences ,0301 basic medicine ,Cell type ,Sucrose ,C4 photosynthesis ,Physiology ,Starch ,Carbohydrate synthesis ,Plant Science ,Biology ,Carbohydrate metabolism ,mesophyll cells ,01 natural sciences ,Carbon Cycle ,03 medical and health sciences ,chemistry.chemical_compound ,Photosynthesis ,Review Papers ,AcademicSubjects/SCI01210 ,Bundle sheath cells ,starch ,sucrose ,Metabolism ,Plants ,Vascular bundle ,Plant Leaves ,030104 developmental biology ,chemistry ,Biochemistry ,010606 plant biology & botany - Abstract
The two-cell type C4 photosynthetic pathway requires both anatomical and biochemical specialization to achieve a functional CO2-concentrating mechanism. While a great deal of research has been done on Kranz anatomy and cell-specific expression and activity of enzymes in the C4 pathway, less attention has been paid to partitioning of carbohydrate synthesis between the cell types of C4 leaves. As early as the 1970s it became apparent that, in the small number of species examined at the time, sucrose was predominantly synthesized in the mesophyll cells and starch in the bundle sheath cells. Here we discuss how this partitioning is achieved in C4 plants and explore whether this is a consequence of C4 metabolism or indeed a requirement for its evolution and efficient operation., C4 plants partition starch and sucrose biosynthesis between bundle sheath and mesophyll cells. Here we review why and how this is achieved, and discuss the relevance for C4 photosynthesis.
- Published
- 2021
47. Expanding the Enzyme Repertoire for Sugar Nucleotide Epimerization: the CDP-Tyvelose 2-Epimerase from Thermodesulfatator atlanticus for Glucose/Mannose Interconversion
- Author
-
Bernd Nidetzky, Christian Rapp, Hansjörg Weber, Stevie Van Overtveldt, Tom Desmet, and Koen Beerens
- Subjects
Carbohydrate synthesis ,specificity ,STRUCTURAL-ANALYSIS ,Mannose ,CARBOHYDRATE EPIMERASES ,Context (language use) ,catalytic mechanism ,01 natural sciences ,Applied Microbiology and Biotechnology ,REDUCTASE ,CDP-glucose ,multistep enzyme catalysis ,UDP-GALACTOSE 4-EPIMERASE ,03 medical and health sciences ,chemistry.chemical_compound ,CRYSTAL-STRUCTURE ,BIOSYNTHESIS ,Nucleotide ,030304 developmental biology ,YERSINIA-PSEUDOTUBERCULOSIS ,chemistry.chemical_classification ,0303 health sciences ,PURIFICATION ,Ecology ,epimerase ,010405 organic chemistry ,Biology and Life Sciences ,Substrate (chemistry) ,sugar nucleotide ,CYTIDINE DIPHOSPHATE 3 ,SUBSTRATE-SPECIFICITY ,0104 chemical sciences ,carbohydrates (lipids) ,Carbohydrate Epimerases ,carbohydrate synthesis ,Enzyme ,chemistry ,Biochemistry ,Earth and Environmental Sciences ,short-chain dehydrogenase/reductase (SDR) ,lipids (amino acids, peptides, and proteins) ,NAD+ kinase ,6-DIDEOXYHEXOSES ,Food Science ,Biotechnology - Abstract
Epimerization of sugar nucleotides is central to the structural diversification of monosaccharide building blocks for cellular biosynthesis. Epimerase applicability to carbohydrate synthesis can be limited, however, by the high degree of substrate specificity exhibited by most sugar nucleotide epimerases. Here, we discovered a promiscuous type of CDP-tyvelose 2-epimerase (TyvE)-like enzyme that promotes C-2 epimerization in all nucleotide (CDP, UDP, GDP, ADP, and TDP)-activated forms of D-glucose. This new epimerase, originating from Thermodesulfatator atlanticus, is a functional homodimer that contains one tightly bound NAD(+)/subunit and shows optimum activity at 70 degrees C and pH 9.5. The enzyme exhibits a k(cat) with CDP-D-glucose of -1.0 min(-1) (pH 7.5 and 60 degrees C). To characterize the epimerase kinetically and probe its substrate specificity, we developed chemoenzymatic synthesis for CDP-D-mannose, CDP-D-deoxy-D-glucose, CDP-3-deoxy-o-glucose, and CDP-6-deoxy-D-xylo-hexopyranos-4-ulose. Attempts to obtain CDP-D-paratose and CDP-D-tyvelose were not successful. Using high-resolution carbohydrate analytics and in situ nuclear magnetic resonance (NMR) to monitor the enzymatic conversions (60 degrees C and pH 7.5), we show that the CDP-D-mannose/CDP-D-glucose ratio at equilibrium is 0.67 (+/- 0.1), determined from the kinetic Haldane relationship and directly from the reaction. We further show that deoxygenation at sugar C-6 enhances the enzyme activity 5-fold compared to CDP-D-glucose, whereas deoxygenation at C-3 renders the substrate inactive. Phylogenetic analysis places the T. atlanticus epimerase into a distinct subgroup within the sugar nucleotide epimerase family of SDRs (short-chain dehydrogenases/reductases), for which the current study now provides functional context. Collectively, our results expand an emerging toolbox of epimerase-catalyzed reactions for sugar nucleotide synthesis. IMPORTANCE Epimerases of the sugar nucleotide-modifying class of enzymes have attracted considerable interest in carbohydrate (bio)chemistry for the mechanistic challenges and the opportunities for synthesis involved in the reactions catalyzed. The discovery of new epimerases with an expanded scope of sugar nucleotide substrates used is important to promote mechanistic inquiry and can facilitate the development of new enzyme applications. Here, a CDP-tyvelose 2-epimerase-like enzyme from Thermodesulfatator atlanticus is shown to catalyze sugar C-2 epimerization in CDP-glucose and other nucleotide-activated forms of D-glucose. The reactions are new to nature in the context of enzymatic sugar nucleotide modification. The current study explores the substrate scope of the discovered C-2 epimerase and, based on modeling, suggests structure-function relationships that may be important for specificity and catalysis.
- Published
- 2021
- Full Text
- View/download PDF
48. To develop optimized protocols for automated glycan assembly using the Glyconeer synthesizer
- Author
-
Gupta, Tanistha
- Subjects
Automation ,500 Natural sciences and mathematics::540 Chemistry and allied sciences::547 Organic chemistry ,Glyconeer ,Carbohydrate Synthesis - Abstract
The introduction of automation in carbohydrate synthesis has paved the way towards getting faster access to complex, immunologically relevant glycans. Automated assembly enables the synthesis of oligomers and polymers (as long as 50 mers) to be accomplished in weeks, compared to months or even years when performed via conventional solution phase synthesis. However, the optimization of the entire process still lies in its infancy, and standard protocols for the automated synthesis of oligosaccharides are not yet available. The work in this thesis aims to develop optimized protocols for the automated assembly of complex glycans using the Glyconeer synthesizer, which was accomplished by firstly, optimizing the synthesis of building blocks. Since building block synthesis is very often the most laborious and time consuming step in the entire process of automation, ready availability of standard protocols for procuring multi-gram quantities of these monomers further reduces the time required to synthesize complex glycans via automated synthesis. Thereafter, with multi-gram quantities of building blocks in hand, the automated synthesis of glycans using the Glyconeer synthesizer were streamlined. Optimized protocols for automated glycan assembly of complex glycans were established by streamlining the variables associated with the glycosylation reaction, namely, temperature, number of glycosylation cycles and concentration of donor (in this case building blocks). Therefore, the AGA of Lewisx epitope was optimized to establish optimal glycosylation conditions for the building blocks (Chapter 3) on a 0.0125 mmol scale. With these optimized protocols, the synthesis was then scaled up to a 0.025 mmol scale to procure milligram quantities of the Lex epitope. Secondly, in order to establish the reproducibility of these protocols, the optimized synthetic protocols were applied to procure a library of Lewis antigens, namely, Lewisa, Lewisy and Lewisb antigens (Chapter 4). The successful synthesis of these antigens further confirmed the reproducibility of the established synthetic protocols. Thereafter, these synthetic protocols were then validated for synthesizing longer structures, which was achieved by synthesizing a library of poly-N-acetyl glucosamine oligomers, such as, tetramer, pentamer and hexamer (Chapter 5). The oligomers synthesized were then used to determine the substrate specificity of the glucosamine hydrolase from xii Pseudomonas aeruginosa. Finally, the boundaries of AGA were further pushed by synthesizing one of the most difficult linkage, namely, the beta mannose linkage via automation (Chapter 6), which makes the synthesis of complex N-glycans possible by automated synthesis. In a nutshell, the work performed in this thesis aims to overcome the challenge of gaining quick access to pure oligosaccharides by establishing standard protocols for synthesizing immunologically relevant glycans via automated glycan assembly, thereby significantly reducing the time required to synthesize these glycans.
- Published
- 2021
- Full Text
- View/download PDF
49. Expanding the Enzyme Repertoire for Sugar Nucleotide Epimerization: The CDP-Tyvelose 2-Epimerase from
- Author
-
Christian, Rapp, Stevie, van Overtveldt, Koen, Beerens, Hansjörg, Weber, Tom, Desmet, and Bernd, Nidetzky
- Subjects
CDP-glucose ,multistep enzyme catalysis ,carbohydrate synthesis ,epimerase ,short-chain dehydrogenase/reductase (SDR) ,specificity ,Enzymology and Protein Engineering ,catalytic mechanism ,sugar nucleotide - Abstract
Epimerases of the sugar nucleotide-modifying class of enzymes have attracted considerable interest in carbohydrate (bio)chemistry for the mechanistic challenges and the opportunities for synthesis involved in the reactions catalyzed. The discovery of new epimerases with an expanded scope of sugar nucleotide substrates used is important to promote mechanistic inquiry and can facilitate the development of new enzyme applications., Epimerization of sugar nucleotides is central to the structural diversification of monosaccharide building blocks for cellular biosynthesis. Epimerase applicability to carbohydrate synthesis can be limited, however, by the high degree of substrate specificity exhibited by most sugar nucleotide epimerases. Here, we discovered a promiscuous type of CDP-tyvelose 2-epimerase (TyvE)-like enzyme that promotes C-2 epimerization in all nucleotide (CDP, UDP, GDP, ADP, and TDP)-activated forms of d-glucose. This new epimerase, originating from Thermodesulfatator atlanticus, is a functional homodimer that contains one tightly bound NAD+/subunit and shows optimum activity at 70°C and pH 9.5. The enzyme exhibits a kcat with CDP-d-glucose of ∼1.0 min−1 (pH 7.5 and 60°C). To characterize the epimerase kinetically and probe its substrate specificity, we developed chemoenzymatic synthesis for CDP-d-mannose, CDP-6-deoxy-d-glucose, CDP-3-deoxy-d-glucose, and CDP-6-deoxy-d-xylo-hexopyranos-4-ulose. Attempts to obtain CDP-d-paratose and CDP-d-tyvelose were not successful. Using high-resolution carbohydrate analytics and in situ nuclear magnetic resonance (NMR) to monitor the enzymatic conversions (60°C and pH 7.5), we show that the CDP-d-mannose/CDP-d-glucose ratio at equilibrium is 0.67 (±0.1), determined from the kinetic Haldane relationship and directly from the reaction. We further show that deoxygenation at sugar C-6 enhances the enzyme activity 5-fold compared to CDP-d-glucose, whereas deoxygenation at C-3 renders the substrate inactive. Phylogenetic analysis places the T. atlanticus epimerase into a distinct subgroup within the sugar nucleotide epimerase family of SDRs (short-chain dehydrogenases/reductases), for which the current study now provides functional context. Collectively, our results expand an emerging toolbox of epimerase-catalyzed reactions for sugar nucleotide synthesis. IMPORTANCE Epimerases of the sugar nucleotide-modifying class of enzymes have attracted considerable interest in carbohydrate (bio)chemistry for the mechanistic challenges and the opportunities for synthesis involved in the reactions catalyzed. The discovery of new epimerases with an expanded scope of sugar nucleotide substrates used is important to promote mechanistic inquiry and can facilitate the development of new enzyme applications. Here, a CDP-tyvelose 2-epimerase-like enzyme from Thermodesulfatator atlanticus is shown to catalyze sugar C-2 epimerization in CDP-glucose and other nucleotide-activated forms of d-glucose. The reactions are new to nature in the context of enzymatic sugar nucleotide modification. The current study explores the substrate scope of the discovered C-2 epimerase and, based on modeling, suggests structure-function relationships that may be important for specificity and catalysis.
- Published
- 2020
50. RNA-Seq analysis reveals the growth and photosynthetic responses of rapeseed (Brassica napus L.) under red and blue LEDs with supplemental yellow, green, or white light
- Author
-
Xuyang Yao, Yixuan Wen, Zheng Chen, Liu Xiaoying, Haifeng Ding, Shirong Guo, Mohammad Shah Jahan, and Xu Zhigang
- Subjects
0106 biological sciences ,0301 basic medicine ,Rapeseed ,Antioxidant ,medicine.medical_treatment ,Plant physiology ,Brassica ,Carbohydrate synthesis ,RNA-Seq ,Plant Science ,Horticulture ,Photosynthesis ,01 natural sciences ,Biochemistry ,Article ,law.invention ,03 medical and health sciences ,law ,Genetics ,medicine ,Light responses ,Secondary metabolism ,biology ,biology.organism_classification ,030104 developmental biology ,010606 plant biology & botany ,Biotechnology ,Light-emitting diode - Abstract
Compound light is required for plant growth and development, but the response mechanisms of plants are undercharacterized and not fully understood. The present study was undertaken to evaluate the effect of supplemental light (green light, G; white light, W; yellow light, Y) added to red–blue light (RB) and sole W on the growth and photosynthesis of rapeseed seedlings. The results revealed that supplemental G/W improved the growth and photosynthesis of seedlings, but supplemental Y significantly reduced the photosynthetic rate and palisade tissue layer. Sole W caused similar responses in terms of growth, leaf development, oxidative damage, and antioxidant capability as supplemental Y. In total, 449, 367, 813, and 751 differentially expressed genes (DEGs) were identified under supplemental G, Y, and W and sole W, respectively, compared to RB. The DEGs under different lights were closely associated with pathways such as light stimulus and high-light response, root growth, leaf development, photosynthesis, photosynthesis-antenna proteins, carbohydrate synthesis and degradation, secondary metabolism, plant hormones, and antioxidant capacity, which contributed to the distinct growth and photosynthesis under different treatments. Our results suggest that Y is more likely substituted by other wavelengths to achieve certain effects similar to those of supplemental Y, while G has a more distinctive effect on rapeseed. Taken together, supplementation RB with G/W promotes the growth of rapeseed seedlings in a controlled environment.
- Published
- 2020
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